Genetic and Biochemical Characterization of the Pseudoalteromonas tetraodonis Alkaline (cid:1) -Carrageenase

An alkaline (cid:1) -carrageenase, Cgk-K142, was found in the culture broth of a deep-sea bacterium, Pseudoalteromonas tetraodonis JAM-K142. A gene for the enzyme was cloned and expressed. Purified recombinant Cgk-K142 (rCgk-K142) showed an optimal pH of about 8.8 in glycine-NaOH buffer at 30 (cid:1) C and of about 8.0 in MOPS buffer at 50 (cid:1) C. The optimal temperature for the enzyme was 55 (cid:1) C at pH 8.0. rCgk-K142 was unstable, but (cid:2) - and (cid:3) -carrageenans, non-degradative substrate homologs, extensively enhanced its stability. The nucleotide sequence of the gene for Cgk-K142 comprised 1,194 bp, and the deduced amino acid sequence (397 amino acids) showed a high level of similarity to the (cid:1) -carrageenase of P. carrageenovora , with 94% identity. Another gene for a (cid:1) -carrageenase-like protein was found downstream of the gene for Cgk-K142. The nucleotide sequence of that gene consisted of 966 bp (321 amino acids), and it showed the highest similarity, at 64% identity, to protein CgkB of P. carrageenovora , which has been reported as an incomplete 57-amino acid sequence.